The ble resistance gene as a new selectable marker for Trypanosoma brucei: fly transmission of stable procyclic transformants to produce antibiotic resistant bloodstream forms

D Jefferies, P Tebabi, D Le Ray, E Pays

Research output: Contribution to journalA1: Web of Science-article

Abstract

We describe here the stable transformation of Trypanosoma brucei using a new selectable marker for kinetoplastid protozoa, the Sh ble, or phleomycin, resistance gene. A plasmid containing this gene targeted to the tubulin gene locus by homologous sequences was introduced into procyclic trypanosomes by electroporation and cells selected for antibiotic resistance. Southern analysis of stable transformants showed that the plasmid had been integrated into the tubulin locus by homologous recombination. Analysis of bloodstream stage transformants, produced by transmission through the vector Glossina, showed that the resistance gene was conserved and expressed in these forms in the absence of selective drug pressure. In both procyclic and bloodstream forms, transcription of the ble gene appears to originate from the upstream tubulin promoter, despite the presence of a VSG promoter in the integrated construct. The generation of stable bloodstream transformants for the first time will facilitate the study of gene function and expression during the trypanosome life cycle, and aid in the investigation of genetic exchange in these organisms
Original languageEnglish
JournalNucleic Acids Research
Volume21
Pages (from-to)191-195
ISSN0305-1048
Publication statusPublished - 1993

Keywords

  • B780-tropical-medicine
  • Protozoology
  • Trypanosoma brucei brucei
  • Genetics
  • Drug resistance
  • Transmission
  • Laboratory medicine
  • Techniques
  • Blotting

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